Isolated plant virus promoters are useful in the genetic engineering of plants to produce transgenic plants with desired phenotypic characteristics. In order to produce such transgenic plants, an isolated promoter is inserted into a vector and operably linked to a heterologous DNA sequence. Plant cells can then be transformed in a variety of ways by DNA constructs containing an isolated promoter fused to heterologous DNA sequences. The result of this transformation is that the promoter operably linked to the heterologous DNA is inserted into the genome of the transformed plant cell. Furthermore, the regulation of the expression of the heterologous DNA in the transformed plant cell is controlled by the promoter.
There are a variety of different approaches for producing a desired phenotype in a transgenic plant, depending on the nature of the heterologous sequences coupled to the isolated promoter. For example, expression of a novel gene that is not normally expressed in plant or in a particular tissue of a plant may confer a phenotypic change. Alternatively, the expression of a sense or an antisense construct introduced into transgenic plants can cause the inhibition of expression of endogenous plant genes. This inhibition of expression can, in turn, produce the desired phenotypic change.
There is a need for a variety of different promoters to be used in the genetic engineering of plants. These promoters are of several types. Constitutive promoters are one such commonly used type of promoter. Constitutive promoters are those which are capable of expressing operably linked DNA sequences in all tissues of a plant throughout normal development. In contrast to constitutive promoters, tissue-specific promoters are those promoters that are capable of selectively expressing heterologous DNA sequences in certain plant tissues. Promoters may also be inducible, e.g., by application of external inducing agents. Constitutive, inducible and tissue-specific promoters are used in the genetic engineering of plants, and have value for many different potential applications in this field.
Constitutive plant promoters may be obtained by isolating the regulatory region of a plant operon that is constitutively expressed. In addition to promoters obtained from plant genes, there are also promoters of bacterial and viral origin which have been used to constitutively express novel sequences in plant tissues. Examples of such promoters from bacteria include the octopine synthase (ocs) promoter, the nopaline synthase (nos) promoter and others derived from native Ti plasmids (see Herrera-Estrella et al, Nature, 303:209–213, 1983). The 35S and 19S promoters of cauliflower mosaic virus are commonly used examples of viral promoters, (see Odel et al, Nature, 313:810–812, 1985).
In contrast to constitutive promoters, tissue-specific promoters are generally isolated from the promoter regions of plant genes that are selectively expressed in a specific plant tissue. These promoters can be fused with a heterologous DNA sequence and used to transform a plant cell to create transgenic plants that selectively express the heterologous DNA in a specific tissue. For example, the promoter regions from the fruit-specific, ethylene regulated genes E4 and E8 and from the fruit-specific polygalacturonase gene have been used to direct fruit specific expression of a heterologous DNA sequence in transgenic tomato plants. (See Cordes et al, Plant Cell, 1;1025–1034, 1989; Deikman et al, EMBO J., 7;3315–3320, 1988; and Della Penna et al, Proc. Natl. Acad. Sci. USA, 83:6420–6424, 1986.)
Aspects of characterization, including genomic cloning, molecular characterization and sequencing, and description of the promoters, from several different plant viruses have been described, including cauliflower mosaic virus (CaMV), Hull, in “Virus Taxonomy”, eds. Murphy et al., Wein, New York, Springer-Verlag, p 189–192, 1995; commelina yellow mottle virus (ComYMV), Medberry et al, Nuc. Acid Res., 18:5505–5513, 1990; rice tungo bacilliform virus (RTBV), Hay et al, Nuc. Acids Res., 19:2615–2621, 1991; sugarcane bacilliform virus (ScBV), Bouhida et al, J. Gen. Virol., 74:15–22, 1993; soybean chlorotic mottle virus (SbCMV) Hasegawa et al, Nuc. Acids Res., 17:9993–10013, 1989; figwort mosaic virus (FMV), Richins et al, Nuc. Acids Res., 15:8451–8466, 1987; carnation etch ringspot virus (CERV), Hull et al, EMBO J., 5:3083–3090, 1986; peanut chlorotic streak virus (PCSV), Reddy et al, Phytopathol., 83:129–133, 1993; strawberry vein banding virus (SVBV), GeneBank Accession No. X97304; and cacao swollen shoot virus (CSSV), GeneBank Accession No. L14546.
Cassava vein mosaic virus (previously referred to as CVMV, now referred to as CsVMV) was described by Calvert et al, J. Gen. Virol., 76:1271–1276, 1995, and sequence data is also published as GeneBank Accession Nos. U59751 and U20341. In addition, the CsVMV promoter was recently described by Verdaguer et al, Plant Mol. Biol., 31:1129–1139, 1996.
The discovery of both new constitutive promoters and new tissue-specific promoters is desired for the controlled expression of various nucleic acid sequences that are engineered into transgenic plants. There are many valuable potential applications of genetic engineering of plants. A variety of plant promoters with different characteristics and which are effective in different species of plants is desirable in order to bring these potential applications into practice.